Chronic alcohol consumption is associated with tolerance to ethanol. Cell membranes isolated from ethanol-fed rodents are resistant to many of the effects of alcohol on membrane structure and function. However, the mechanisms of the effects of ethanol on biological membranes and the acquired tolerance are largely unknown. The hypothesis, that ethanol effects on membrane functions is the result of modulation of the membrane dielectric constant and that tolerance results from attenuation of this effect, shall be tested. This hypothesis is consistent with the suggestion that tolerance is associated with reduced partition of ethanol to specific membrane domains. Synaptosomes and synaptic plasma membranes will be isolated from ethanol-fed rats and their pair-fed controls. The effects of alcohols on the membrane dielectric constant in membrane preparations from ethanol-fed rats and their controls will be estimated by three independent methods. The polarity sensing probes Prodan and Laurdan will be used to estimate the modulation by alcohols of the membrane polarity at the membrane surface and at the core from the fluorescence emission spectra of these probes. Fluorescent surface pH and surface potential indicators, as well as EPR surface potential probes, will be used to evaluate the effect of alcohols on the surface dielectric constant from the apparent pKa of the pH indicators and surface acidic groups. The modulation of the membrane permeability of lipophilic ions by alcohols will be evaluated in terms of modulation of the membrane dielectric constant. Selective quenching of fluorescent and EPR probes on the exofacial membrane leaflet will be used to evaluated the dielectric constant and its modulations by alcohols on each of the membrane surfaces. Fractionation of lipid components of these membranes and reconstitution will be used to evaluate the contribution of specific membrane components to the acute and chronic effects of alcohol on the membrane dielectric constant. These studies should lead to a better understanding of the molecular mechanisms of the effect of ethanol on membranes function and the acquired tolerance observed in alcoholics.